The present invention relates polymeric hydrogel blends, and more particularly to polymeric hydrogel blends that exhibit angiogenic and vascularizing activity.
Hydrogels are usually formed by chemical and physical cross-linking reaction between molecules using for example light energy and light-activated free radical initiators to promote chemical cross-linking (U.S. Pat. No. 4,957,744 to della Valle). Photo cross-linkable polymers are typically chemically modified (activated) to provide reactive groups for the crosslinking reaction. Successful examples include photocurable hydrogels made from synthetic materials, such as polyethylene glycol (Sawhney, et al., Bioerodible Hydrogels Based On Photopolymerized Poly(Ethylene Glycol)-Co-Poly(A-Hydroxy Acid) Diacrylate Macromers, Macromolecules, 26(4):581-587 (1993)), and also from macromolecules of biological origin. Hydrogels made of derivatized polysaccharides are formed using such a procedure (U.S. Pat. No. 5,763,504 to Takehisa Matsuda, et al.).
In the field of medical research, hydrogels are used to provide scaffolds to support cell growth in tissue replacement and regeneration applications, or to serve as drug delivery vehicles by adding drugs to the hydrogel. The drug may either be entrapped in the gel, or the molecules can be ionically or covalently bound to the backbone of the hydrogel (U.S. Pat. No. 5,677,276 to Dickerson, et al.). Hydrogels can also be implanted without the addition of cells or drugs, and serve some space filling cosmetic or anatomical purposes, such as tissue augmentation (Marler, et al., Soft-Tissue Augmentation With Injectable Alginate And Syngeneic Fibroblasts, Plast Reconstr Surg, 105(6):2049-58 (2000)). Bioactive hydrogels intrinsically elicit a specific host response, such as promoting the migration of certain cells or tissue ingrowth (Agren, M S., An Amorphous Hydrogel Enhances Epithelialisation Of Wounds, Acta Derm Venereol, 78(2):119-22 (1998)).
Host reactions of interest, angiogenesis and neovascularization, entail the formation of new blood vessels sprouting from the vascular bed surrounding the hydrogel implant and their growth into the hydrogel implant, respectively. Several angiogenic substances have been identified, including small fragments of the tissue matrix polysaccharide hyaluronate, ranging between 4 and 25 disaccharides in length (D C West, et al., Angiogenesis Induced By Degradation Products Of Hyaluronic Acid, Science, 228(4705):1324-6 (1985)). Hyaluronate, a tissue extracellular matrix glycosaminoglycan of approximately 1 million Daltons molecular weight, is naturally broken down into smaller fragments by the action of an enzyme, hyaluronidase, or by hydrolysis. Fractionation of hyaluronate may be accomplished in vitro using the enzyme hyaluronidase (Turner, et al., Self-Association Of Hyaluronate Segments In Aqueous NaCl Solution, Arch. Biochem. Biophys., 265(2):484-95 (1988)). However, to isolate the desired fragment size, time and manipulation are required.
In view of the art, there is a need for angiogenic hyaluronate-containing hydrogels that avoids the time-consuming and costly fractionation methods associated with the production of small hyaluronate fragments. Accordingly, it is an object to provide angiogenic hyaluronate-containing hydrogels that avoids these time-consuming and costly production methods.
The present invention provides hydrogels having angiogenic activity and pre-gel blends for synthesizing the hydrogels. The pre-gel blend includes a mixture of a polysaccharide material at least partially substituted with an unsaturated, cross-linking moiety, and a non-angiogenic hyaluronic acid or salt thereof. Preferably, the polysaccharide material has an average molecular weight of at least about 10,000 Daltons, with at least about 40,000 Daltons being more preferred. Classes of polysaccharide material to be used are starch or starch derivatives, water-soluble gums, or mixtures thereof. Examples of starch or starch derivatives are dextrans, curdlans, succinoglycans, pullulans, cellulose derivatives, cyclodextrins, or mixtures thereof Examples of water-soluble gum are alginates, carageenens, xanthans, galactomannans, or mixtures thereof. In a particular embodiment, the starch or starch derivative is dextran. Preferably, the unsaturated, cross-linking moiety is selected from the group consisting of acrylates, esters, ethers, thioethers, amides, enamides, sulfonyl esters or mixtures thereof, with acrylate being particularly preferred. The non-angiogenic hyaluronic acid or salt preferably has an average molecular weight of at least about 500,000 Daltons, with at least about 1,000,000 Daltons being more preferable.
The hydrogels of the present invention are a reaction product prepared by a process which includes: admixing the polysaccharide material that is at least partially substituted with the unsaturated, cross-linking moiety, and the non-angiogenic hyaluronic acid or salt thereof, with a free-radical initiator in a solvent; and cross-linking the mixture to form the hydrogel. The free-radical initiator is either a chemical initiator and a non-chemical initiator, with a non-chemical initiator (e.g., UV initiator) being preferred. A preferred class of UV initiators are acetophenone derivatives. The solvent is preferably water, a water-based solvent, a water-miscible organic solvent, or mixtures thereof. The other components are as described above.
The present invention also provides kits for preparing the hydrogels. In one embodiment the kit includes: a first container containing a mixture of the partially substituted, polysaccharide material and the non-angiogenic hyaluronic acid or salt thereof; and a second container containing a free-radical initiator. In another embodiment the kit includes: a first container containing the partially substituted, polysaccharide material; a second container containing the non-angiogenic hyaluronic acid or salt thereof; and a third container containing the free-radical initiator. Preferably, the kits further include an instruction pamphlet for preparing the hydrogel.
Advantageously, the pre-gel blends and hydrogels of the present invention omit the use of angiogenic hyaluronate fragments as a starting material while providing angiogenic hyaluronate-containing hydrogels. These and other advantages of the invention will become more readily apparent from the detailed description set forth below.